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three-dimensional and multidimensional microscopy: image acquisition and processing XXX 2023

作者： Ghezzi, Alberto Farina, Andrea Lenz, Armin J.M. Soldevila, Fernando Tajahuerce, Enrique Bassi, Andrea Valentini, Gianluca D’Andrea, Cosimo Politecnico di Milano Dipartimento di Fisica Piazza L. da Vinci 32 Milano20133 Italy Consiglio Nazionale delle Ricerche IFN Piazza L. da Vinci 32 Milan20133 Italy Universitat Jaume I Avda. Sos Baynat s/n Castelló12071 Spain UMR 8552 Collège de France 24 rue Lhomond Paris75005 France Center for NanoScience and Technology @PoliMi Istituto Italiano di Tecnologia via Pascoli 70/3 via Pascoli 70/3 Milan20133 Italy

ISBN: (纸本)9781510658752

Time-resolved multispectral fluorescence microscopy provides a 4D hypercube dataset with high specificity for cellular examination. However, this is generally obtained by significantly increasing the measurement time, which is quite limiting for in vivo measurement or with photosensitive sample. It is possible to reduce the measurement effort with a novel microscopy framework exploiting Compressive Sensing based on Single-Pixel Camera. In this work, we present a compressive sensing system and validate it with a cellular sample. Data fusion with a high-resolution camera image allows us to tackle the well-known problem of low-resolution in single-pixel imaging. © 2023 SPIE. All rights reserved.

关键词： Compressed sensing

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MAGNIFY: molecule anchorable gel-enabled nanoscale in-situ fluorescence microscopy for nanoscale imaging of biomolecules 30

MAGNIFY: molecule anchorable gel-enabled nanoscale in-situ f...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXX 2023

作者： Klimas, Aleksandra Gallagher, Brendan R. DiBernardo, Emma Cheng, Zhangyu Zhao, Yongxin Department of Biological Sciences Carnegie Mellon University PittsburghPA United States

ISBN: (纸本)9781510658752

Expansion microscopy (ExM) is a powerful imaging strategy that offers a low-cost solution for interrogating biological systems at the nanoscale using conventional optical microscopes. It achieves this by physically and isotropically magnifying preserved biological specimens embedded in a cross-linked water-swellable hydrogel. However, most reported techniques are unable to preserve endogenous epitopes due to strong protease digestion used to expand samples. In addition, these protocols rely on mechanically fragile hydrogels that only expand by at most 4.5× linearly. We present a new ExM framework, Molecule Anchorable Gel-enabled Nanoscale In-situ Fluorescence microscopy (MAGNIFY), that exhibits a broad retention of nucleic acids, proteins, and lipids without the need for a separate anchoring step. By using a mechanically sturdy hydrogel, MAGNIFY is capable of expanding biological specimens up to 11×. This facilitates nanoscale imaging (~25-nm effective resolution) using an ∼280-nm diffraction-limited objective lens on a conventional optical microscope and can be furthered to ~15 nm effective resolution if combined with computational methods such as Super-resolution Optical Fluctuation Imaging (SOFI). Here, we demonstrate that MAGNIFY provides a generalized solution for imaging nanoscale subcellular features of a broad range of biological specimens. We also show that MAGNIFY provides a novel, accessible tool for improving the precision, utility, and generality of nanoscopy. © 2023 SPIE. All rights reserved.

关键词： Fluorescence microscopy

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Automatic detection of alignment errors in cryo-electron tomography

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JOURNAL OF STRUCTURAL BIOLOGY 2025年第1期217卷 108153页

作者： de Isidro-Gomez, F. P. Vilas, J. L. Carazo, J. M. Sorzano, C. O. S. Campus Univ Autonoma Ctr Nacl Biotecnol CNB CSIC Biocomp Unit Darwin 3 Madrid 28049 Spain Univ Autonoma Madrid Madrid 28049 Spain

Cryo-electron tomography is an imaging technique that allows the study of the three-dimensional structure of a wide range of biological samples, from entire cellular environments to purified specimens. This technique collects a series of images from different views of the specimen by tilting the sample stage in the microscope. Subsequently, this information is combined into a three-dimensional reconstruction. To obtain reliable representations of the specimen of study, it is mandatory to define the acquisition geometry accurately. This is achieved by aligning all tilt images to a standard reference scheme. Errors in this step introduce artifacts into the final reconstructed tomograms, leading to loss of resolution and making them unsuitable for detailed sample analysis. This publication presents algorithms for automatically assessing the alignment quality of the tilt series and their classification based on the residual errors provided by the alignment algorithms. If no alignment information is available, a set of algorithms for calculating the residual vectors focused on fiducial markers is also presented. This software is accessible as part of the Xmipp software package and the Scipion framework.

关键词： Cryo-electron tomography Tilt-series alignment Electron tomography Electron microscopy image processing Structural biology

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Two plus one is almost three: a fast approximation for multi-view deconvolution

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BIOMEDICAL OPTICS EXPRESS 2022年第1期13卷 147-158页

作者： Huepfel, Manuel Merino, Manuel Fernandez Bennemann, Johannes Takamiya, Masanari Rastegar, Sepand Tursch, Anja Holstein, Thomas W. Nienhaus, G. Ulrich Karlsruhe Inst Technol KIT Inst Appl Phys Wolfgang Gaede Str 1 D-76131 Karlsruhe Germany Karlsruhe Inst Technol KIT Inst Biol & Chem Syst IBC D-76021 Eggenstein Leopoldshaftn Germany Heidelberg Univ Ctr Organismal Studies COS D-69120 Heidelberg Germany Karlsruhe Inst Technol KIT Inst Nanotechnol D-76021 Eggenstein Leopoldshafen Germany Univ Illinois Dept Phys Urbana IL 61801 USA

Multi-view deconvolution is a powerful image-processing tool for light sheet fluorescence microscopy, providing isotropic resolution and enhancing the image content. However, performing these calculations on large datasets is computationally demanding and time-consuming even on high-end workstations. Especially in long-time measurements on developing animals, huge amounts of image data are acquired. To keep them manageable, redundancies should be removed right after image acquisition. To this end, we report a fast approximation to three-dimensional multi-view deconvolution, denoted 2D+1D multi-view deconvolution, which is able to keep up with the data flow. It first operates on the two dimensions perpendicular and subsequently on the one parallel to the rotation axis, exploiting the rotational symmetry of the point spread function along the rotation axis. We validated our algorithm and evaluated it quantitatively against two-dimensional and three-dimensional multi-view deconvolution using simulated and real image data. 2D+1D multi-view deconvolution takes similar computation time but performs markedly better than the two-dimensional approximation only. Therefore, it will be most useful for image processing in time-critical applications, where the full 3D multi-view deconvolution cannot keep up with the data flow. Published by The Optical Society under the terms of the Creative Commons Attribution 4.0 License.

关键词： Beam shaping image fusion image processing image quality image resolution Optical transfer functions

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Broadening Circular and Linear SAR image and 3D volume parameters for accurate localization 29

Broadening Circular and Linear SAR image and 3D volume param...

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Conference on Algorithms for Synthetic Aperture Radar imagery xxix

作者： Pepin, Matthew USAF Albuquerque NM USA

ISBN: (纸本)9781510650671;9781510650664

Determining accurate location with Synthetic Aperture Radar (SAR) images is hindered not only by the limited resolution of the image but also by the non-rectangular nature of the process that produces the image, and the presence of multiple returns from different height objects at the same range. three dimensional SAR volume measurements can differentiate different height scatterers but imaging methods are limited by the measurement scenario. Interpolation of images and volumes can normalize the collected data but can add inaccuracy to pixel and voxel impulse response (IPR) and localization estimates. Regions of best achievable accuracy define the imaging algorithm performance envelope. Ideal localization occurs at the estimation bound of modeled point scatterers, and is also indicated by pixel and voxel size or IPR main lobe width. The performance envelope of SAR measurement scenarios, radar parameters, and imaging and estimation algorithms can refine system parameters of accurate localization. Use of analogous Circular and Linear SAR (CSAR and LSAR) scenarios enables joint trade-off of flight path along with imaging parameters. An ideal spherical construct underlies joint CSAR and LSAR, two and three dimensional SAR scenarios. This construct also ties CSAR and LSAR imaging to spherical ray-trace creation of phase history. For different scenarios defined through a spherical model of parameters, localization performance of the imaging envelope, IPR and estimation accuracy are examined. Along with sample size and ideal chirp waveform sampling, the azimuth and elevation aperture is varied to include ideal and sub-sampling, and extension to achieve un-aliased images and volumes. Methods expanding the envelope are investigated.

关键词： Imaging arrays Synthetic aperture radar 3D modeling 3D image processing Convolution Solid modeling Radar 3D acquisition image resolution

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3D super-resolution live- cell imaging with radial symmetry and Fourier light-field microscopy

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BIOMEDICAL OPTICS EXPRESS 2022年第11期13卷 5574-5584页

作者： Han, Keyi Hua, Xuanwen Vasani, Vishwa Kim, Ge-Ah R. Liu, Wenhao Takayama, Shuichi Jia, Shu Georgia Inst Technol Wallace H Coulter Dept Biomed Engn Atlanta GA 30332 USA Emory Univ Atlanta GA 30332 USA Georgia Inst Technol George W Woodruff Sch Mech Engn Atlanta GA 30332 USA Georgia Inst Technol Sch Mat Sci & Engn Atlanta GA 30332 USA Georgia Inst Technol Parker H Petit Inst Bioengn & Biosci Atlanta GA 30332 USA

Live-cell imaging reveals the phenotypes and mechanisms of cellular function and their dysfunction that underscore cell physiology, development, and pathology. Here, we report a 3D super-resolution live-cell microscopy method by integrating radiality analysis and Fourier light-field microscopy (rad-FLFM). We demonstrated the method using various live-cell specimens, including actins in Hela cells, microtubules in mammary organoid cells, and peroxisomes in COS-7 cells. Compared with conventional wide-field microscopy, rad-FLFM realizes scanning-free, volumetric 3D live-cell imaging with sub-diffraction-limited resolution of similar to 150 nm (x-y) and 300 nm (z), milliseconds volume acquisition time, six-fold extended depth of focus of similar to 6 mu m, and low photodamage. The method provides a promising avenue to explore spatiotemporal-challenging subcellular processes in a wide range of cell biological research.

关键词： image processing image quality Imaging techniques Live cell imaging Plenoptic imaging three dimensional imaging

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DOF adjustment and 3D free viewpoint viewing of Z stack data of fluorescent microscope using image processing 28

DOF adjustment and 3D free viewpoint viewing of Z stack data...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXVIII 2021

作者： Hagiwara, Tsuneyuki 3rd Research Section Optical Research Laboratory Research and Development Division Nikon Corporation Japan

ISBN: (数字)9781510641341

ISBN: (纸本)9781510641334

In recent years, confocal fluorescence microscopes have become widespread and many Z-stack data have been collected. However, it is the current situation that there is hardly a method for three-dimensional observation of huge amounts of data with sufficient resolution and without loss of information. In light of this situation, we have developed an image processing algorithm, referred to as arbitrary blur algorithm (ABA), which enables post-adjustment of DOF and 3D image observation from a free viewpoint, with high resolution and without loss of information. In this report, we first explain the principle of image processing that enables depth of field (DOF) adjustment and 3D image observation from a free viewpoint using Z-stack data. Next, the results of the simulation are introduced. In the simulation, we show the comparison result of the point image by MIP (maximum intensity projection) method and the point image of the proposed method and clarify the superiority of the proposed method. Next, we introduce the results of the experimental verification. For experimental verification, we show the results using wide field fluorescence microscope Z-stack data and confocal fluorescence microscope Z-stack data. As confocal data, observation examples of mouse hippocampal neuron synapses are shown in comparison with the results displayed by existing software. © 2021 Copyright SPIE.

关键词： Microscopes

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three-dimensional and multidimensional microscopy: image acquisition and processing XXVIII

Three-Dimensional and Multidimensional Microscopy: Image Acq...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXVIII 2021

ISBN: (纸本)9781510641334

The proceedings contain 15 papers. The topics discussed include: sparsity-based approach for 3D super-resolution microscopy from correlation information of high emitter-density frames;quantitative phase imaging-based machine learning approaches for the analysis of adherent and suspended cells;automatic nuclei segmentation based on fuzzy C- mean;evaluation of different illumination designs for tunable 3D structured illumination microscopy through model-based restoration;high-throughput 3D imaging with compressive lens-free tomography;intensity uniformity optimization in spatial-light-modulator-based multifocal microscope;and method for measurement of collagen monomer orientation in fluorescence microscopy.

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Robust gaussian fitting for super-resolution microscopy with fast parameters initial estimation 28

Robust gaussian fitting for super-resolution microscopy with...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXVIII 2021

作者： Qin, Shun Chan, Wai Kin Tsinghua Shenzhen International Graduate School Shenzhen China

ISBN: (数字)9781510641341

ISBN: (纸本)9781510641334

Efficient accurate Gaussian fitting is an important topic in many applications, take localization based super-resolution microscopy and image scanning microscopy for example, which requires localizing plenty of Gaussian patterns for accurately reconstructing a super-resolution image. Existing Gaussian fitting methods usually require inputting a good initial value for all parameters for efficient and robust fitting, which apparently is not suitable for the task of large scale Gaussian fitting with a computer. It would be even more challenge to estimate an appropriate initial value for all parameters and guarantee the robustness of the fitting algorithm for low signal-to-noise ratio measured data with strong background. In this paper, we propose a two-step fitting algorithm for robust and accurate 2D Gaussian fitting without inputting any initial parameter value. Our simulation shows that the performance of the fitting algorithm can be improved significantly with the proposed parameter initial value estimation algorithm. © 2021 Copyright SPIE.

关键词： Optical resolving power

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Sparsity-based approach for 3D super-resolution microscopy from correlation information of high emitter-density frames 28

Sparsity-based approach for 3D super-resolution microscopy f...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXVIII 2021

作者： Dardikman-Yoffe, Gili Eldar, Yonina C. Faculty of Mathematics and Computer Science Weizmann Institute of Science Israel

ISBN: (数字)9781510641341

ISBN: (纸本)9781510641334

Breakthroughs in the field of chemistry have enabled surpassing the classical optical diffraction limit by utilizing photo-activated fluorescent molecules. In the single-molecule localization microscopy (SMLM) approach, a sequence of diffraction-limited images, produced by a sparse set of emitting fluorophores with minimally overlapping point-spread functions is acquired, allowing the emitters to be localized with high precision by simple post-processing. However, the low emitter density concept requires lengthy imaging times to achieve full coverage of the imaged specimen on the one hand, and minimal overlap on the other. Thus, this concept in its classical form has low temporal resolution, limiting its application to slow-changing specimens. In recent years, a variety of approaches have been suggested to reduce imaging times by allowing the use of higher emitter densities. One of these methods is the sparsity-based approach for super-resolution microscopy from correlation information of high emitter-density frames, dubbed SPARCOM, which utilizes sparsity in the correlation domain while assuming that the blinking emitters are uncorrelated over time and space, yielding both high temporal and spatial resolution. However, SPARCOM has only been formulated for the two-dimensional setting, where the sample is assumed to be an infinitely thin single-layer, and thus is unsuitable to most biological specimens. In this work, we present an extension of SPARCOM to the more challenging three-dimensional scenario, where we recover a volume from a set of recorded frames, rather than an image. © 2021 Copyright SPIE.

关键词： Optical transfer function

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